High (5X) 5-methyltetrahydrofolate (MTHF) vs folic acid (FA) pregnancy diets on hypothalamic gene expression of Wistar rat dams upon birth.

This study compared the effects of 5X requirements of FA (10mg/kg) vs equimolar 5-methyletetrahydrofolate (MTHF, 10.4mg/kg) provided during pregnancy on hypothalamic (arcuate nucleus of the hypothalamus) gene expression profiles of Wistar rat dams upon birth. Methods: Whole-transcriptome analysis by RNA-sequencing was performed with Illumina HiSeq 2500 instrument (paired-end 2x125 bp sequencing at a depth of ˜20 million paired reads per sample). Sequence reads that passed quality filters were aligned to the rat genome using STAR aligner and differential gene expression was analyzed by DESeq2 using a padj <0.05. SYBR Green assay was used to validate candidate genes via qRT–PCR. Results: A total of 279 genes were identified to be differentially expressed genes in the hypothalamus of dams at birth fed high (5X) MTHF compared to 5X-FA diets during pregnancy, with 170 up-regulated and 109 down-regulated genes. Using goseq for pathway enrichment analysis, we identified several enriched pathways that associated with the long-term phenotype of the dam including the KEGG neuroactive-ligand receptor interaction pathway. Conclusion: This study provides novel insight towards the differential effects of folate form during pregnancy on regulating dynamic changes in gene expression of the maternal brain upon birth. The results of our study may led to hypothesis testing to identify novel candidate genes and pathways targeted by folate during pregnancy that have a role in disease on-set and/or prevention. Overall design: Hypothalamic (arcuate nucelus of the hypothalamus) mRNA profiles from Wistar rat dams immediately after giving birth.

本研究比较了妊娠期间饲喂5倍剂量叶酸（folic acid，FA）与等摩尔剂量5-甲基四氢叶酸（5-methyletetrahydrofolate，MTHF）对分娩后Wistar大鼠母鼠下丘脑弓状核基因表达谱的影响。方法：采用Illumina HiSeq 2500测序仪开展RNA测序的全转录组分析，测序模式为双端2×125 bp测序，每个样本的测序深度约为2000万条配对读段。将通过质量过滤的序列读段通过STAR比对工具比对至大鼠基因组，使用DESeq2进行差异基因表达分析，筛选阈值为校正后P值（padj）<0.05。采用SYBR Green分析法通过qRT-PCR对候选基因进行验证。结果：与妊娠期间饲喂5倍剂量FA日粮的母鼠相比，分娩后饲喂5倍剂量MTHF日粮的母鼠下丘脑中共鉴定出279个差异表达基因，其中170个基因上调、109个基因下调。通过goseq进行通路富集分析，我们发现多个与母鼠长期表型相关的富集通路，包括京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes，KEGG）神经活性配体-受体相互作用通路。结论：本研究为妊娠期间不同叶酸形式对分娩后母鼠脑组织基因表达的动态变化的调控差异提供了新的见解。本研究结果可为后续假设验证提供依据，以鉴定妊娠期间叶酸靶向调控的、与疾病发生和/或预防相关的新型候选基因及通路。整体设计：分娩后即刻采集的Wistar大鼠母鼠下丘脑弓状核的mRNA表达谱。